Re: manuscript submission entitled: High Yield of Culture-based Diagnosis in a TBendemic Setting
Reviewers’ comments and authors’ replies
1) Reviewer : Susan S Dorman
In this prospective cohort study conducted in a routine TB program setting in Cape
Town from 2002-2003, the authors enrolled pulmonary TB suspects who then
underwent routine program-clinician assessments with the addition of mycobacterial
culture to sputum testing (otherwise smear microscopy). The authors determined the
relative and absolute incremental diagnostic yields of culture, and the number of
cultures needed to diagnose one additional TB case. These metrics are uncommonly
used in TB and contribute to the novelty and importance of this manuscript. An
important weakness of the study/manuscript is that liquid and solid culture data were
not able to be analyzed separately, since "the individual results for solid and liquid
media were not recorded in the database.'
We agree that this is a weakness of the study as the results of the solid and liquid media
were not reported separately on the laboratory report. We have added this as a
limitation of the study in the discussion.
Major Compulsory Revision
1. Greater than 50% of study participants were excluded from the analyses, and this
represents a weakness of the analytic approach. This was apparently done to ensure
that each analyzed participant had 2 smears and 2 cultures. Two revisions are
requested: analysis of participants having 1 smear and 1 culture, and inclusion of a
schematic showing the flow of participants and reasons for exclusion from analysis.
This is indeed a weakness of the study and so we changed our analysis to calculate
incremental yields by using the expected proportion for the missing specimens based on
the observed proportions of positive smears and cultures, using a method described by
Rieder et al in 2005. We had seen this method before and had decided not to use it but
your useful comment and the use of it in another article recently published in PloS One
on the same subject prompted us to use it. We have presented the data in a clearer way
and have also added a flow diagram for the exclusion of patients.
Minor Essential Revision
1. page 9, lines 199-200: the manuscript data and results do support the use of culture,
but not clearly the use of LIQUID culture.
We have made the correction.
Discretionary Revision
1. Lines 183-184: the analytical specificity of smear is not high, in that morphology
cannot distinguish between mycobacterial species. However, the authors are clearly
correct that smear does have excellent PPV in high incidence areas. Perhaps this
sentence should be rephrased as simply "Although smear is less sensitive than culture,
nevertheless it has excellent PPV in high incidence areas."
We have used your suggestion and made the change in the text.
2) Reviewer : Scott Heysell
a) Major revisions:
Given that the study period is more remote than expected (2002-2003) it warrants
explanation of why these data, instead of more current investigation, were used.
To be honest, there is no single reason we are submitting data compiled around 2005
for publication in 2012. A few things occurred: 1) We published two other papers, one
on cross-contamination. We considered the lab quality paper, published in 2012, critical
to any papers on the substantive data; 2) The first and corresponding author live in
Montreal, so some of the details of the dataset took some time to obtain. Demers has
recently travelled to South Africa to work with the local investigators in order to iron out
these details. As a result of this, we delayed submission of this manuscript until we
were confident in the data.
Finally, while the study period is a few years ago, we do not see how this affects the
qualtity of the data and the inferences about diagnostic yield. We are aware of the
introduction of XPert as a novel diagnostic platform since our study began, which is why
we noted this in our manuscript.
Given that other studies have demonstrated an improved yield of culture over smear
and that it is, as mentioned, a WHO endorsed goal for TB programs, the relevance of a
study of this kind would be to describe how many patients were started on TB
treatment based on the use of culture over smear. It is mentioned in the methods that
TB treatment episodes were obtained from notes, TB register etc. For instance, if
subjects were lost to follow-up before culture results were known, this is a limitation
to scale-up. Or if smear negative subjects with a high suspicion of TB were started on
empiric TB treatment prior to culture results then
this diminishes the programmatic benefit of culture.
We agree that this is an important follow-up aim. We previously included all the lab
data and the patient treatment data in a long draft of this manuscript, but there were
too many tables and the manuscript lacked focus. Therefore, our proximal goal is to
describe the laboratory yield of culture, when compared to smear. Our subsequent goal
will be to write a manuscript on the treatment initiation as a function of the diagnostic
modality.
Furthermore, a major advantage to culture is drug-susceptibility testing. Susceptibility
results should be mentioned. For which patients/samples was DST performed? If the
treatment episode is known, did DST change the treatment course or facilitate
referral?
Thank you, this is an excellent point. We have added the information on susceptibility in
the article. However, the numbers of MDR were very low, in this cohort, and we could
not analyse this any further.
Line 157, the statement regarding the absolute yield of two smears appears
inconsistent with table 2. Taking the SNP to mean smear positive on the second
specimen, then 100 of the 369 smear positives were diagnosed in this way, of which
84 were culture positive? Which is >5.7% absolute yield? I see where this is
represented in table 2, and translates to a NND of 18 for performing a second smear.
Perhaps table 1a and table 2 are all that is necessary with elimination of table 1b?
We apologize for any confusion in the manner we have computed and presented this in
the previous version of the manuscript. We have recalculated and presented the results
in a new way (cf Methods and Results for more details). Hopefully things will be clearer
this time.
Line 165, the HIV VCT is low for confirmed TB infection, while not out of range for
certain endemic areas this highlights the incredible need for integrated HIV/TB
services and warrants comment as to whether this proportion has changed currently
with availability of antiretrovirals. The proportion is low enough to where conclusions
regarding HIV may be misinformed.
This is an important point and we have addressed it in the results and discussion.
Line 179, suggest tempering the “remarkable” assessment of the results, and as
mentioned before, the lack of routine HIV testing limits the reliability of these findings
in that subset. Thus suggest stating that “these findings demonstrate that in a
programmatic setting culture is of expectedly higher yield, requiring 9 additional
specimens to detect one additional case of TB, and in HIV infected patients, the
number of additional specimens may be as low as 3.”
Agree, we have made the suggested change.
Major limitations to implementation are cost and laboratory training/expertise rather
than lack of knowledge that culture will increase yield. A comment addressing the
anticipated cost of scaling up laboratory services compared to theoretical missed
cases of TB will be important.
We have added a note regarding the cost of scaling up lab services in the discussion.
The paragraph on comparison to Xpert and other rapid molecular technologies is
important but the sensitivity and specificity, as mentioned, is known in various
settings. It may be preferable to discuss the advantages and disadvantages of the
molecular technologies compared to culture with respect to the total number of
patients started on treatment, the time-to-treatment initiation, and the time-totreatment initiation with an active regimen based on DST.
This is an interesting point, but as noted above, our dataset predates Xpert, so a
detailed discussion contrasting molecular vs. culture-based diagnosis would be largely a
theoretical discussion, not informed by the data from this study. We prefer our
discussion to be directly pertinent to the study we describe and not use the discussion
as a platform for extrapolation to other diagnostic modalities for which we have no
data.
b) Minor revisions:
Line 68 typo, “however”
The typo was corrected.
Line 96, will need red font color changed to black
The font color was corrected.
Line 124-127 is confusing. I take this to read scanty smears were considered positive?
If so, then can simply state eliminate reference to prior WHO definition and previous
published work.
We have simplified the text in that regard.
Line 133, were NTM + cultures actually excluded from analysis? Should these not have
been considered as SPCN? Or smear positive, culture not TB? In other settings without
culture these patients would be treated as having TB. How would this have influenced
yield? It does not appear necessary to re-analyze these data, but rather comment
within the discussion.
We have added more information on NTM. However, we have decided to remove them
from the analysis as they only were found in 30 SE.
Line 143, by what statistical method was HIV influence analyzed? This line is a little
vague.
This was indeed vague and a correction was made.
Line 190, reference mislabeled?
We are not sure of which reference but we have verified all references.
Line 208, AY and RY are introduced for the first time in the text and should be spelled
out for clarification.
The correction was made
Line 239, DST is used for the first time in the text and should be spelled out for
clarification.
The correction was made
c) Discretionary:
Line 62 redundant, could just say “However, smear microscopy has a detection limit of
about 5000-10 000 organisms/mL [3]- compared to 10-100 organisms/mL for culture
[4]”
Arguably there is some redundancy here, but we consider this a key concept for the
manuscript. All TB patients with < 5000 bacterial / mL of sputum will be classified as
smear-negative in a microscopy-based diagnostic program. In this case, we prefer to be
redundant in order to make this point.